Our studies on the assembly of murine leukemia viruses have involved a correlative morphological and biochemical approach into two problems. First, we studied the role of cellular processes in regulating leukemia virus production. We found that high virus producing cell lines exhibited budding of virus into intracytoplasmic vesicles or vacuoles, as well as the well-characterized budding at the cell surface. Our approach has been to study the inhibition of virus production in these cells by using microtubule depolymerizing agents which presumably would prevent intracytoplasmic vesicles containing virus from arriving at the cell surface. Our preliminary studies showed that there was a 2 1/2 to 5-fold inhibition of virus production when inhibitory agents such as vinblastine or nocadozole were used. Correlative EM studies suggest budding from the surface is maintained with the drugs. The second problem is the elucidation of the specific changes that accompany the conversion of "immature" to "mature" virus particles. We have partially purified the proteolytic factor that cleaves Pr659a9, the precursor polyprotein to the internal core proteins, and found that when it was added back to immature cores they were apparently converted to mature cores. However, quantitation of the complex events of this process at a biochemical level have not yet been done. These require isolation of large amounts of the murine Pr659a9 proteolytic factor. Due to the fact that the factor is present in small amounts and extremely unstable, we have had to devise new methods for purifying it. We are currently trying affinity and other column chromatographic methods in an effort to obtain large enough amounts of the factor for such in vitro studies.